Methods of treating generalized pustular psoriasis with an antagonist of CCR6 or CXCR2

ABSTRACT

The present disclosure provides, inter alia, methods of treating generalized pustular psoriasis (GPP) by administering an effective amount of a Chemokine Receptor 6 (CCR6) antagonist and/or a C-X-C motif chemokine receptor 2 (CXCR2) antagonist. Also provided herein are methods of modulating dysregulated IL-36 signaling in a subject in need thereof and methods of reducing neutrophil, inflammatory dendritic cell (iDC), and/or CD4 T cell accumulation in a subject in need thereof, said methods, include dministering an effective amount of a Chemokine Receptor 6 (CCR6) antagonist and/or a C-X-C motif chemokine receptor 2 (CXCR2) antagonist. In some embodiments, the CCR6 and/or CXCR2 antagonist has the formula:

CROSS-REFERENCES TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.16/241,469 filed Jan. 7, 2019, which application claims the benefit ofpriority under 35 U.S.C § 119(e) to U.S. Provisional Application Ser.No. 62/614,927 filed Jan. 8, 2018 and U.S. Provisional Application Ser.No. 62/715,503 filed Aug. 7, 2018, the disclosures of each of which areincorporated herein by reference in their entirety.

STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSOREDRESEARCH AND DEVELOPMENT

Not Applicable

REFERENCE TO A “SEQUENCE LISTING,” A TABLE, ORA COMPUTER PROGRAM LISTINGAPPENDIX SUBMITTED ON A COMPACT DISK

Not Applicable

BACKGROUND

Generalized pustular psoriasis (GPP) is a rare disease for which thereis a dearth of clinical research and no universally acceptedevidence-based guidelines for its treatment and management (Benjegerdeset al. Psoriasis (Auckl) 2016; 6:131-44.). Biologic therapies that areeffective in the more common plaque form of psoriasis are ineffective inGPP (Benjegerdes et al. Psoriasis (Auckl) 2016; 6:131-44. Mansouri etal. Expert Opin Biol Ther 2013; 13(12):1715-30.), and much neededtreatments that directly target GPP have not been developed (Mahil etal. Semin Immunopathol 2016; 38(1):11-27. Navarini et al. J Eur AcadDermatol Venereol 2017; 31(11):1792-9. Robinson et al. J Am AcadDermatol 2012; 67(2):279-88.

The present disclosure addresses the need for promising therapies thattarget and ameliorate GPP symptoms and provides related advantages aswell.

BRIEF SUMMARY OF THE INVENTION

The present disclosure provides methods of treating generalized pustularpsoriasis (GPP), palmo-plantar psoriasis (PPP), acute generalizedexanthematous pustulosis (AGEP), hydradenitis suppurativa (HS),dermatitis herpetiformis, and/or pemphigus vulgaris said methodcomprising administering an effective amount of an antagonist ofChemokine Receptor 6 (CCR6) and/or C-X-C motif chemokine receptor 2(CXCR2).

In another aspect, provided herein are methods of modulatingdysregulated IL-36 signaling in a subject in need thereof, said methodcomprising administering to the subject an effective amount of anantagonist of Chemokine Receptor 6 (CCR6) and/or C-X-C motif chemokinereceptor 2 (CXCR2).

In a further aspect, provided herein are methods of reducing neutrophil,inflammatory dendritic cell (iDC), and/or CD4 T cell accumulation in asubject in need thereof comprising administering to the subject aneffective amount of an antagonist of Chemokine Receptor 6 (CCR6) and/orC-X-C motif chemokine receptor 2 (CXCR2).

In some embodiments, the CCR6 and/or CXCR2 antagonist has the formula:

where each variable is described below.

In some embodiments, the CCR6 and/or CXCR2 antagonist has the formula:

where each variable is described below.

In some embodiments, the CCR6 and/or CXCR2 antagonist is a compoundshown in FIG. 1 .

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.129:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.123:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.136:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.138:

or a pharmaceutically acceptable salt thereof.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A-1AJ provides particular structures for compounds describedherein.

FIG. 2A-D shows the gating scheme used to identify iDC, neutrophils andCD4⁺αβ T cells isolated from IL-36-treated skin as discussed in FIG. 3 ,FIG. 5 , and FIG. 10 . The cells were first gated on the Live, CD45⁺population, followed by the Thy-1 vs CD11b gating (Panel A). The cellscircled in the lower left corner were gated on Ly6C vs Ly6G to identifyiDCs and neutrophils (Panel B). The circled cells in the upper leftcorner of panel A were then gated on TCRαβ vs TCRγδ (Panel C). The cellscirculated in panel C were gated on CD8α vs CD4 to identify CD4⁺αβ Tcells (Panel D).

FIG. 3A-D shows the markedly different inflammatory cell skininfiltrates generated in the Imiquimod model of Plaque Psoriasis and theIL-36 Model of GPP. Panels A and B show the number of cells per gramisolated from mouse skin after four daily treatments with IMQ (A, greybars with horizontal stripes) or IL-36α (B, grey bars with diagonalstripes). Black bars indicate the number of cells per gram isolated fromcontrol-treated skin, topical Vaseline (VAS) for the imiquimodexperiments (A) and intradermal PBS for the IL-36 experiments (B). PanelC displays relative representation of leukocyte subsets withinIL-36-treated versus imiquimod-treated skin: comparison among individualexperiments. The percentage of T cells, neutrophils and iDC wascalculated for the total live CD45⁺ infiltrate for 5 individualimiquimod experiments and 7 individual IL-36 experiments, eachexperiment utilizing at least 5 individual mice. The mean of the means(and SEM) for these experiments is shown. Panel D shows that percent ofT cells that expressed the indicated immunophenotype isolated from theskin of mice after treatment with imiquimod (grey bars with horizontalstripes) or IL-36α (grey bars with diagonal stripes). Each bar indicatesthe mean and SEM of ten mice from a single experiment, representative ofat least 5 repeats. All populations shown were first gated as live(AQUA-live/dead negative) and CD45⁺. For panels a and b, T cells weregated as Thy-1⁺/CD11b⁻ cells expressing either TCRβ or TCRγδ. Ly6G⁺ andLy6C^(hi)/Ly6G⁻ cells were gated within the Thy-1⁻/CD11b⁺ population.For panel c, each of the indicated populations was calculated as percentof the total Thy-1⁺/CD11b⁻ population expressing either TCRβ or TCRγδ.

FIG. 4A-B shows multiplex analysis of CCL20 and CXCL2 proteinsdemonstrates significant increase of both proteins after 4 dailyintradermal injections of IL-36α. CCL20 protein levels are plotted inpanel A; CXCL2 protein levels are plotted in panel B.

FIG. 5A-C shows expression of CCR6 and CXCR2 by leukocytes accumulatingin skin in response to intradermal IL-36α injections. Cells isolatedfrom IL-36-treated ears of 20 mice were pooled and stained withunconjugated specific MAb (as indicated above each column) orisotype-matched control, followed by second stage MAb using standardprocedures. Unbound second stage was blocked with normal mouse, rat andhamster serum, followed by directly labeled monoclonal antibodies.Gating for each cell type is indicated to the left of each row: Myeloidcells are panel A; Neutrophils are panel B; and CD4 T Cells are panel C.Percent of cells brighter than the isotype-matched control is indicatedwithin the flow cytometry plot if greater than 5%. Staining of pooledcells is representative of 4 repeat experiments.

FIG. 6A-B shows that Compound 1.136 ameliorates inflammatory swelling ofIL-36α-injected ears. Panel A plots ear thickness of mice dosed dailywith Compound 1.136 at the indicated doses (or with α-IL-17RA) duringthe IL-36α-induced GPP model. Ear thickness was measured by caliperafter 4 days of treatment. Panel B Time course of ear thickness for theexperiment shown in (A), comparing the 90 mg/kg dose of compound 1.136to α-IL-17RA. Ten mice per data point. Statistics from Mann-Whitney rankorder test. Note: titration experiments showed the effects of α-IL-17RAto plateau at 200 μg per mouse per day, and the mice in this experimentwere dosed at 500 μg per mouse per day (data not shown). n.s., notsignificant p<0.05*,p<0.0005***,p<0.0001****.

FIG. 7A-B shows ear thickness data. Panel A shows a time course fortitration of Compound 1.136 shown in FIG. 6A. Panel B showsisotype-matched control for anti-IL-17RA treatment of IL-36α-inflamedskin. Ears of five mice in each group were inflamed by daily intradermalinjections of PBS or activated IL-36α as described in the text. Somegroups also received daily intraperitoneal injections of 500 μg/mouse ofanti-IL-17RA or 500 μg/mouse of a rat IgG2a isotype-matched negativecontrol for the anti-IL-17RA MAb. The Mann-Whitney Rank Order testestablished significance between the effects of anti-IL-17RA MAb and itsisotype matched control on days 3, 4 and 5. *p<0.05, **p<0.01. Althoughthe effects of anti-IL17RA reached saturation at the 200 μg/mouse/daydose (compare to 500 μg/mouse/day in FIG. 3 ), anti-IL-17RA is comparedto its isotype-matched control at 500 mg/kg/day to demonstrate thateffects were not non-specific effects of the isotype even at these veryhigh levels.

FIG. 8A-D shows that Compound 1.136 substantially improves histology ofIL-36α-injected ears. Ears were acquired from sacrificed mice after 4days of 90 mg/kg IL-36α or PBS treatment (Panel A is PBS+Vehicle; PanelB is IL-36+Vehicle). During treatment, mice were dosed daily with 1%HPMC (the vehicle for Compound 1.136; Panels A & B), with Compound 1.136in vehicle (Panel C) or with anti-IL17RA (Panel D). Ears were fixed andembedded using standard FFPE techniques, sectioned and stained usingstandard hematoxylin and eosin (H&E) staining techniques. Sections shownare representative of at least five different sections from fivedifferent ears.

FIG. 9A-B shows that Compound 1.136 substantially reduces epidermalthickness of ears injected with activated IL-36α. Panel A, top row showsa section of an entire width of ear after 4 daily IL-36α injections.Panel A, second row shows a higher magnification of the top imagefocusing on the lesional area. Panel A, rows three and four showlesional areas from Compound 1.136-treated IL-36α-treated mouse ear andmouse ear injected with PBS instead of IL-36α. Black bars indicate wherethe each of the 7 individual epidermal thickness measurements were takenfor all sections graphed in Panel B. Panel B, ear thickness measurementsfrom 8 mice from each treatment group. Each dot represents the mean of 7epidermal thickness measurements from the ear of 1 mouse. Ears wereacquired from sacrificed mice after 4 daily treatments of activatedIL-36α (or PBS). During treatment, mice were dosed daily subcutaneouslywith 1% HPMC (the vehicle) with Compound 1.136 at 90 mg/kg/day s.c. invehicle, or with anti-IL17RA at 200 μg/mouse/day IP in PBS. Ears werefixed and embedded using standard FFPE techniques, sectioned and stainedusing standard hematoxylin and eosin (H&E) staining techniques. Sectionsshown are representative of at least five different sections from eightdifferent ears. Statistics from Mann-Whitney rank order test p<0.05*.

FIG. 10A-C shows that Compound 1.136 significantly reduces accumulationof CD4 T cells (Panel A), neutrophils (Panel B), and inflammatorydendritic cells (Panel C) within IL-36-treated skin. Ears were acquiredfrom sacrificed mice after 4 daily IL-36α (or PBS control) treatments.IL-36α-injected mice received vehicle alone, Compound 1.136 (90mg/kg/day s.c. on the back) or α-IL-17RA (200 μg/mouse/day in theperitoneum). Statistical analysis by Mann-Whitney rank-order test. Oneexperiment is shown with 10 mice per group and is representative of 3repeats. n.s., not significant, p<0.005**, p<0.0001****.

FIG. 11A-E shows the characterization of Ly6C^(hi) myeloid cellsaccumulating in skin after intradermal IL-36α injections. Cells wereisolated from 20 ears after four daily IL-36α injections, then pooledand stained with unconjugated specific MAb (as indicated within eachpanel, light grey curves) or isotype-matched controls (dark greycurves), followed by an anti-Ig second stage polyclonal Ab usingstandard procedures. Unbound second stage was blocked with normal mouse,rat and hamster serum, followed by directly conjugated MAbs. Panel Auses a CD103 specific MAb; Panel B uses a Flt3 specific MAb; Panel Cuses a CD205 specific MAb; Panel D uses a CD11c specific MAb; and PanelE uses a F4/80 specific MAb. Myeloid cells were gated as in FIG. 5A.Staining of pooled cells shown is representative of 3 repeatexperiments.

DETAILED DESCRIPTION OF THE INVENTION

General

Generalized pustular psoriasis (GPP) is a rare inflammatory skindisorder with an etiology distinct from the more common plaquepsoriasis. GPP patients often do not respond to therapeutic agentstypically used for plaque psoriasis. Antagonists of CCR6 and/or CXCR2including the compounds disclosed herein have been previously shown toameliorate inflammation in a model of plaque psoriasis. Surprisingly,the present disclosure demonstrates that an antagonist of CCR6 and/orCXCR2 can be used to effectively treat generalized pustular psoriasis(GPP). In addition to treating GPP, related diseases such aspalmo-plantar psoriasis (PPP), acute generalized exanthematouspustulosis (AGEP), hydradenitis suppurativa (HS), dermatitisherpetiformis, and pemphigus vulgaris can also be treated using themethods described herein.

Chemokine directed therapy is designed to block the migration ofinflammatory leukocytes into tissues from the peripheral blood, thuspreventing them from participating in and amplifying any existingautoimmune lesions, thereby allowing the inflammatory cytokineenvironment to dissipate. Genetic studies demonstrate that GPP isstrongly associated with dysfunctions in the IL-36 cytokine axis, andmany aspects of GPP can be re-created in the mouse by intradermalinjection of pre-activated IL-36α cytokine. The present disclosuredemonstrates that the immune cells infiltrating IL-36α-injected mouseskin are of dramatically different composition than those infiltratingimiquimod- (IMQ-) treated skin, an accepted model of plaque psoriasis inBalb/c mice. The findings disclosed herein suggest that CCR6 and CXCR2antagonists may constitute a novel target class for a mechanisticallydistinct therapeutic approach to treat GPP as well as related PPP, AGEP,HS, dermatitis herpetiformis, and pemphigus vulgaris diseases.

Abbreviations and Definitions

Unless otherwise indicated, the following terms are intended to have themeaning set forth below. Other terms are defined elsewhere throughoutthe specification.

The term “alkyl”, by itself or as part of another substituent, means,unless otherwise stated, a straight or branched chain hydrocarbonradical, having the number of carbon atoms designated (i.e. C₁₋₈ meansone to eight carbons). Examples of alkyl groups include methyl, ethyl,n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, n-pentyl,n-hexyl, n-heptyl, n-octyl, and the like.

The term “cycloalkyl” refers to hydrocarbon rings having the indicatednumber of ring atoms (e.g., C₃₋₆ cycloalkyl) and being fully saturatedor having no more than one double bond between ring vertices.“Cycloalkyl” is also meant to refer to bicyclic and polycyclichydrocarbon rings such as, for example, bicyclo[2.2.1]heptane,bicyclo[2.2.2]octane, etc.

The term “cycloheteroalkyl” refers to a cycloalkyl ring having theindicated number of ring vertices (or members) and having from one tofive heteroatoms selected from N, O, and S, which replace one to five ofthe carbon vertices, and wherein the nitrogen and sulfur atoms areoptionally oxidized, and the nitrogen atom(s) are optionallyquaternized. The cycloheteroalkyl may be a monocyclic, a bicyclic or apolycylic ring system. Non limiting examples of cycloheteroalkyl groupsinclude pyrrolidine, imidazolidine, pyrazolidine, butyrolactam,valerolactam, imidazolidinone, hydantoin, dioxolane, phthalimide,piperidine, 1,4-dioxane, morpholine, thiomorpholine,thiomorpholine-S-oxide, thiomorpholine-S,S-oxide, piperazine, pyran,pyridone, 3-pyrroline, thiopyran, pyrone, tetrahydrofuran,tetrhydrothiophene, quinuclidine, and the like. A cycloheteroalkyl groupcan be attached to the remainder of the molecule through a ring carbonor a heteroatom.

As used herein, a wavy line, “

”, that intersects a single, double or triple bond in any chemicalstructure depicted herein, represent the point attachment of the single,double, or triple bond to the remainder of the molecule. Additionally, abond extending to the center of a ring (e.g., a phenyl ring) is meant toindicate attachment at any of the available ring vertices. One of skillin the art will understand that multiple substituents shown as beingattached to a ring will occupy ring vertices that provide stablecompounds and are otherwise sterically compatible. For a divalentcomponent, a representation is meant to include either orientation(forward or reverse). For example, the group “-C(O)NH-” is meant toinclude a linkage in either orientation: —C(O)NH— or —NHC(O)—, andsimilarly, “—O—CH₂CH₂—” is meant to include both —O—CH₂CH₂— and—CH₂CH₂—O—.

The terms “alkoxy,” “alkylamino” and “alkylthio” (or thioalkoxy) areused in their conventional sense, and refer to those alkyl groupsattached to the remainder of the molecule via an oxygen atom, an aminogroup, or a sulfur atom, respectively. Additionally, for dialkylaminogroups, the alkyl portions can be the same or different and can also becombined to form a 3-7 membered ring with the nitrogen atom to whicheach is attached. Accordingly, a group represented as dialkylamino or—NR^(a)R^(b) is meant to include piperidinyl, pyrrolidinyl, morpholinyl,azetidinyl and the like.

The terms “halo” or “halogen,” by themselves or as part of anothersubstituent, mean, unless otherwise stated, a fluorine, chlorine,bromine, or iodine atom. Additionally, terms such as “haloalkyl,” aremeant to include monohaloalkyl and polyhaloalkyl. For example, the term“C₁₋₄ haloalkyl” is meant to include trifluoromethyl,2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.

The term “aryl” means, unless otherwise stated, a polyunsaturated,typically aromatic, hydrocarbon group which can be a single ring ormultiple rings (up to three rings) which are fused together or linkedcovalently. Non-limiting examples of aryl groups include phenyl,naphthyl and biphenyl.

The term “heteroaryl” refers to aryl groups (or rings) that contain fromone to five heteroatoms selected from N, O, and S, wherein the nitrogenand sulfur atoms are optionally oxidized, and the nitrogen atom(s) areoptionally quaternized. A heteroaryl group can be attached to theremainder of the molecule through a heteroatom. Non-limiting examples ofheteroaryl groups include pyridyl, pyridazinyl, pyrazinyl, pyrimindinyl,triazinyl, quinolinyl, quinoxalinyl, quinazolinyl, cinnolinyl,phthalazinyl, benzotriazinyl, purinyl, benzimidazolyl, benzopyrazolyl,benzotriazolyl, benzisoxazolyl, isobenzofuryl, isoindolyl, indolizinyl,benzotriazinyl, thienopyridinyl, thienopyrimidinyl, pyrazolopyrimidinyl,imidazopyridines, benzothiaxolyl, benzofuranyl, benzothienyl, indolyl,quinolyl, isoquinolyl, isothiazolyl, pyrazolyl, indazolyl, pteridinyl,imidazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiadiazolyl,pyrrolyl, thiazolyl, furyl, thienyl and the like. Substituents for aheteroaryl ring can be selected from the group of acceptablesubstituents described below.

The above terms (e.g., “alkyl,” “aryl” and “heteroaryl”), in someembodiments, will be optionally substituted. Selected substituents foreach type of radical are provided below.

Optional substituents for the alkyl radicals (including those groupsoften referred to as alkylene, alkenyl, alkynyl and cycloalkyl) can be avariety of groups selected from: halogen, —OR′, —NR′R″, —SR′,—SiR′R″R′″, —OC(O)R′, —C(o)R′, —CO₂R′, —CONR′R″, —OC(O)NR′R″,—NR″C(O)R′, —NR′—C(O)NR″R′″, —NR″C(O)₂R′, —NH—C(NH₂)═NH, —NR′C(NH₂)═NH,—NH—C(NH₂)═NR′, —S(O)R′, —S(O)₂R′, —S(O)₂NR′R″, —NR′S(O)₂R″, —CN and—NO₂ in a number ranging from zero to (2 m′+1), where m′ is the totalnumber of carbon atoms in such radical. R′, R″ and R′″ eachindependently refer to hydrogen, unsubstituted C₁₋₈ alkyl, unsubstitutedaryl, aryl substituted with 1-3 halogens, unsubstituted C₁₋₈ alkyl, C₁₋₈alkoxy or C₁₋₈thioalkoxy groups, or unsubstituted aryl-C₁₋₄ alkylgroups. When R′ and R″ are attached to the same nitrogen atom, they canbe combined with the nitrogen atom to form a 3-, 4-, 5-, 6-, or7-membered ring. For example, -NR′R″ is meant to include 1-pyrrolidinyland 4-morpholinyl.

Similarly, optional substituents for the aryl and heteroaryl groups arevaried and are generally selected from: -halogen, —OR′, —OC(O)R′,—NR′R″, —SR′, —R′, —CN, —NO₂, —CO₂R′, —CONR′R″, —C(O)R′, —OC(O)NR′R″,—NR″C(O)R′, —NR″C(O)₂R′, —NR′—C(O)NR″R′″, —NH—C(NH₂)═NH, —NR′C(NH₂)═NH,C(NH₂)═NR′, —S(O)R′, —S(O)₂R′, —S(O)₂NR′R″, —NR′S(O)₂R″, —N₃,perfluoro(C₁-C₄)alkoxy, and perfluoro(C₁-C₄)alkyl, in a number rangingfrom zero to the total number of open valences on the aromatic ringsystem; and where R′, R″ and R′″ are independently selected fromhydrogen, C₁₋₈ alkyl, C₁₋₈haloalkyl, C₃₋₆ cycloalkyl, C₂₋₈ alkenyl andC₂₋₈ alkynyl. Other suitable substituents include each of the above arylsubstituents attached to a ring atom by an alkylene tether of from 1-4carbon atoms.

Two of the substituents on adjacent atoms of the aryl or heteroaryl ringmay optionally be replaced with a substituent of the formula-T-C(O)—(CH₂)_(q)—U-, wherein T and U are independently —NH—, —O—, —CH₂—or a single bond, and q is an integer of from 0 to 2. Alternatively, twoof the substituents on adjacent atoms of the aryl or heteroaryl ring mayoptionally be replaced with a substituent of the formula-A-(CH₂)_(r)-B-, wherein A and B are independently —CH₂—, —O—, —NH—,—S—, —S(O)—, —S(O)₂—, —S(O)₂NR′— or a single bond, and r is an integerof from 1 to 3. One of the single bonds of the new ring so formed mayoptionally be replaced with a double bond. Alternatively, two of thesubstituents on adjacent atoms of the aryl or heteroaryl ring mayoptionally be replaced with a substituent of the formula—(CH₂)_(s)—X—(CH₂)_(t)—, where s and t are independently integers offrom 0 to 3, and X is —O—, —NR′—, —S—, —S(O)—, —S(O)₂—, or —S(O)₂NR′—.The substituent R′ in —NR′— and —S(O)₂NR′— is selected from hydrogen orunsubstituted C₁₋₆ alkyl.

As used herein, the term “heteroatom” is meant to include oxygen (O),nitrogen (N), sulfur (S) and silicon (Si).

When a variable (e. g. , R¹ or R^(a)) occurs more than one time in anycompound or substituent, its definition on each occurrence isindependent of its definition at every other occurrence. Additionally,combinations of substituents and/or variables are permissible only ifsuch combinations result in stable compounds.

The term “pharmaceutically acceptable salts” is meant to include saltsof the active compounds which are prepared with relatively nontoxicacids or bases, depending on the particular substituents found on thecompounds described herein. When compounds of the present inventioncontain relatively acidic functionalities, base addition salts can beobtained by contacting the neutral form of such compounds with asufficient amount of the desired base, either neat or in a suitableinert solvent. Examples of salts derived frompharmaceutically-acceptable inorganic bases include aluminum, ammonium,calcium, copper, ferric, ferrous, lithium, magnesium, manganic,manganous, potassium, sodium, zinc and the like. Salts derived frompharmaceutically-acceptable organic bases include salts of primary,secondary and tertiary amines, including substituted amines, cyclicamines, naturally-occuring amines and the like, such as arginine,betaine, caffeine, choline, N,N′-dibenzylethylenediamine, diethylamine,2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine,glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperidine, polyamine resins,procaine, purines, theobromine, triethylamine, trimethylamine,tripropylamine, tromethamine and the like. When compounds of the presentinvention contain relatively basic functionalities, acid addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired acid, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable acid additionsalts include those derived from inorganic acids like hydrochloric,hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric,monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,monohydrogensulfuric, hydriodic, or phosphorous acids and the like, aswell as the salts derived from relatively nontoxic organic acids likeacetic, propionic, isobutyric, malonic, benzoic, succinic, suberic,fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric,tartaric, methanesulfonic, and the like. Also included are salts ofamino acids such as arginate and the like, and salts of organic acidslike glucuronic or galactunoric acids and the like (see, for example,Berge, S. M., et al, “Pharmaceutical Salts”, Journal of PharmaceuticalScience, 1977, 66, 1-19). Certain specific compounds of the presentinvention contain both basic and acidic functionalities that allow thecompounds to be converted into either base or acid addition salts.

The neutral forms of the compounds may be regenerated by contacting thesalt with a base or acid and isolating the parent compound in theconventional manner. The parent form of the compound differs from thevarious salt forms in certain physical properties, such as solubility inpolar solvents, but otherwise the salts are equivalent to the parentform of the compound for the purposes of the present invention.

In addition to salt forms, the present invention provides compoundswhich are in a prodrug form. Prodrugs of the compounds described hereinare those compounds that readily undergo chemical changes underphysiological conditions to provide the compounds of the presentinvention. Additionally, prodrugs can be converted to the compounds ofthe present invention by chemical or biochemical methods in an ex vivoenvironment. For example, prodrugs can be slowly converted to thecompounds of the present invention when placed in a transdermal patchreservoir with a suitable enzyme or chemical reagent.

Certain compounds of the present invention can exist in unsolvated formsas well as solvated forms, including hydrated forms. In general, thesolvated forms are equivalent to unsolvated forms and are intended to beencompassed within the scope of the present invention. Certain compoundsof the present invention may exist in multiple crystalline or amorphousforms. In general, all physical forms are equivalent for the usescontemplated by the present invention and are intended to be within thescope of the present invention.

Certain compounds of the present invention possess asymmetric carbonatoms (optical centers) or double bonds; the racemates, diastereomers,geometric isomers, regioisomers and individual isomers (e.g., separateenantiomers) are all intended to be encompassed within the scope of thepresent invention. When a stereochemical depiction is shown, it is meantto refer the compound in which one of the isomers is present andsubstantially free of the other isomer. ‘Substantially free of’ anotherisomer indicates at least an 80/20 ratio of the two isomers, morepreferably 90/10, or 95/5 or more. In some embodiments, one of theisomers will be present in an amount of at least 99%.

The compounds of the present invention may also contain unnaturalproportions of atomic isotopes at one or more of the atoms thatconstitute such compounds. Unnatural proportions of an isotope may bedefined as ranging from the amount found in nature to an amountconsisting of 100% of the atom in question. For example, the compoundsmay incorporate radioactive isotopes, such as for example tritium (³H),iodine-125 (¹²⁵I) or carbon-14 (¹⁴C), or non-radioactive isotopes, suchas deuterium (²H) or carbon-13 (¹³C). Such isotopic variations canprovide additional utilities to those described elsewhere within thisapplication. For instance, isotopic variants of the compounds of theinvention may find additional utility, including but not limited to, asdiagnostic and/or imaging reagents, or as cytotoxic/radiotoxictherapeutic agents. Additionally, isotopic variants of the compounds ofthe invention can have altered pharmacokinetic and pharmacodynamiccharacteristics which can contribute to enhanced safety, tolerability orefficacy during treatment. All isotopic variations of the compounds ofthe present invention, whether radioactive or not, are intended to beencompassed within the scope of the present invention.

The terms “patient” or “subject” are used interchangeably to refer to ahuman or a non-human animal (e.g., a mammal).

The terms “administration”, “administer” and the like, as they apply to,for example, a subject, cell, tissue, organ, or biological fluid, referto contact of, for example, an antagonist of CCR6 and/or CXCR2, apharmaceutical composition comprising same, or a diagnostic agent to thesubject, cell, tissue, organ, or biological fluid. In the context of acell, administration includes contact (e.g., in vitro or ex vivo) of areagent to the cell, as well as contact of a reagent to a fluid, wherethe fluid is in contact with the cell.

The terms “treat”, “treating”, treatment” and the like refer to a courseof action (such as administering an antagonist of CCR6 and/or CXCR2, ora pharmaceutical composition comprising same) initiated after a disease,disorder or condition, or a symptom thereof, has been diagnosed,observed, and the like so as to eliminate, reduce, suppress, mitigate,or ameliorate, either temporarily or permanently, at least one of theunderlying causes of a disease, disorder, or condition afflicting asubject, or at least one of the symptoms associated with a disease,disorder, condition afflicting a subject. Thus, treatment includesinhibiting (e.g., arresting the development or further development ofthe disease, disorder or condition or clinical symptoms associationtherewith) an active disease.

The term “in need of treatment” as used herein refers to a judgment madeby a physician or other caregiver that a subject requires or willbenefit from treatment. This judgment is made based on a variety offactors that are in the realm of the physician's or caregiver'sexpertise.

The terms “prevent”, “preventing”, “prevention” and the like refer to acourse of action (such as administering an antagonist of CCR6 and/orCXCR2, or a pharmaceutical composition comprising same) initiated in amanner (e.g., prior to the onset of a disease, disorder, condition orsymptom thereof) so as to prevent, suppress, inhibit or reduce, eithertemporarily or permanently, a subject's risk of developing a disease,disorder, condition or the like (as determined by, for example, theabsence of clinical symptoms) or delaying the onset thereof, generallyin the context of a subject predisposed to having a particular disease,disorder or condition. In certain instances, the terms also refer toslowing the progression of the disease, disorder or condition orinhibiting progression thereof to a harmful or otherwise undesiredstate.

The term “in need of prevention” as used herein refers to a judgmentmade by a physician or other caregiver that a subject requires or willbenefit from preventative care. This judgment is made based on a varietyof factors that are in the realm of a physician's or caregiver'sexpertise.

The phrase “therapeutically effective amount” refers to theadministration of an agent to a subject, either alone or as part of apharmaceutical composition and either in a single dose or as part of aseries of doses, in an amount capable of having any detectable, positiveeffect on any symptom, aspect, or characteristic of a disease, disorderor condition when administered to the subject. The therapeuticallyeffective amount can be ascertained by measuring relevant physiologicaleffects, and it can be adjusted in connection with the dosing regimenand diagnostic analysis of the subject's condition, and the like. By wayof example, measurement of the serum level of an antagonist of CCR6and/or CXCR2 (or, e.g., a metabolite thereof) at a particular timepost-administration may be indicative of whether a therapeuticallyeffective amount has been used.

The phrase “in a sufficient amount to effect a change” means that thereis a detectable difference between a level of an indicator measuredbefore (e.g., a baseline level) and after administration of a particulartherapy. Indicators include any objective parameter (e.g., serumconcentration) or subjective parameter (e.g., a subject's feeling ofwell-being).

The term “small molecules” refers to chemical compounds having amolecular weight that is less than about 10 kDa, less than about 2 kDa,or less than about 1 kDa. Small molecules include, but are not limitedto, inorganic molecules, organic molecules, organic molecules containingan inorganic component, molecules comprising a radioactive atom, andsynthetic molecules. Therapeutically, a small molecule may be morepermeable to cells, less susceptible to degradation, and less likely toelicit an immune response than large molecules.

The terms “inhibitors” and “antagonists”, or “activators” and “agonists”refer to inhibitory or activating molecules, respectively, for example,for the activation of, e.g., a ligand, receptor, cofactor, gene, cell,tissue, or organ. Inhibitors are molecules that decrease, block,prevent, delay activation, inactivate, desensitize, or down-regulate,e.g., a gene, protein, ligand, receptor, or cell. Activators aremolecules that increase, activate, facilitate, enhance activation,sensitize, or up-regulate, e.g., a gene, protein, ligand, receptor, orcell. An inhibitor may also be defined as a molecule that reduces,blocks, or inactivates a constitutive activity. An “agonist” is amolecule that interacts with a target to cause or promote an increase inthe activation of the target. An “antagonist” is a molecule that opposesthe action(s) of an agonist. An antagonist prevents, reduces, inhibits,or neutralizes the activity of an agonist, and an antagonist can alsoprevent, inhibit, or reduce constitutive activity of a target, e.g., atarget receptor, even where there is no identified agonist.

The terms “modulate”, “modulation” and the like refer to the ability ofa molecule (e.g., an activator or an inhibitor) to increase or decreasethe function or activity of CCR6 and/or CXCR2, either directly orindirectly. A modulator may act alone, or it may use a cofactor, e.g., aprotein, metal ion, or small molecule.

The “activity” of a molecule may describe or refer to the binding of themolecule to a receptor; to catalytic activity; to the ability tostimulate gene expression or cell signaling, differentiation, ormaturation; to antigenic activity; to the modulation of activities ofother molecules; and the like.

As used herein, “comparable”, “comparable activity”, “activitycomparable to”, “comparable effect”, “effect comparable to”, and thelike are relative terms that can be viewed quantitatively and/orqualitatively. The meaning of the terms is frequently dependent on thecontext in which they are used. By way of example, two agents that bothactivate a receptor can be viewed as having a comparable effect from aqualitative perspective, but the two agents can be viewed as lacking acomparable effect from a quantitative perspective if one agent is onlyable to achieve 20% of the activity of the other agent as determined inan art-accepted assay (e.g., a dose-response assay) or in anart-accepted animal model. When comparing one result to another result(e.g., one result to a reference standard), “comparable” frequently(though not always) means that one result deviates from a referencestandard by less than 35%, by less than 30%, by less than 25%, by lessthan 20%, by less than 15%, by less than 10%, by less than 7%, by lessthan 5%, by less than 4%, by less than 3%, by less than 2%, or by lessthan 1%. In particular embodiments, one result is comparable to areference standard if it deviates by less than 15%, by less than 10%, orby less than 5% from the reference standard. By way of example, but notlimitation, the activity or effect may refer to efficacy, stability,solubility, or immunogenicity.

“Substantially pure” indicates that a component makes up greater thanabout 50% of the total content of the composition, and typically greaterthan about 60% of the total content of the composition. More typically,“substantially pure” refers to compositions in which at least 75%, atleast 85%, at least 90% or more of the total composition is thecomponent of interest. In some cases, the component of interest willmake up greater than about 90%, or greater than about 95% of the totalcontent of the composition.

DETAILED DESCRIPTION OF EMBODIMENTS

Methods of Use

Provided herein are methods of using antagonist of Chemokine Receptor 6(CCR6) and/or C-X-C motif chemokine receptor 2 (CXCR2) to prevent,reduce, or maintain leukocyte accumulation (such as neutrophil,inflammatory dendritic cell (iDC), and/or CD4 T cell accumulation),manage and modulate diseases related to IL-36 dysregulation, and thetreatment of diseases such as generalized pustular psoriasis (GPP),palmo-plantar psoriasis (PPP), acute generalized exanthematouspustulosis (AGEP), hydradenitis suppurativa (HS), dermatitisherpetiformis, and pemphigus vulgaris. As described herein, the presentdisclosure demonstrates that antagonist of Chemokine Receptor 6 (CCR6)and/or C-X-C motif chemokine receptor 2 (CXCR2) effectively modulatesleukocyte migration typically observed in subjects experiencing IL-36dysregulation. Administration of a Chemokine Receptor 6 (CCR6) and/orC-X-C motif chemokine receptor 2 (CXCR2) effectively amelioratesinflammation in these subjects.

As such, in one aspect, the present disclosure provides methods oftreating a disease or condition selected from generalized pustularpsoriasis (GPP), palmo-plantar psoriasis (PPP), acute generalizedexanthematous pustulosis (AGEP), hydradenitis suppurativa (HS),dermatitis herpetiformis, or pemphigus vulgaris, said method comprisingadministering an effective amount of an antagonist of Chemokine Receptor6 (CCR6) and/or C-X-C motif chemokine receptor 2 (CXCR2).

In some embodiments, the disease or condition is generalized pustularpsoriasis (GPP). In some embodiments, the disease or condition ispalmo-plantar psoriasis (PPP). In some embodiments, the disease orcondition is acute generalized exanthematous pustulosis (AGEP). In someembodiments, the disease or condition is hydradenitis suppurativa (HS).In some embodiments, the disease or condition is dermatitisherpetiformis. In some embodiments, the disease or condition ispemphigus vulgaris.

In another aspect, provided herein are methods of modulatingdysregulated IL-36 signaling in a subject in need thereof, said methodcomprising administering to the subject an effective amount of anantagonist of Chemokine Receptor 6 (CCR6) and/or C-X-C motif chemokinereceptor 2 (CXCR2).

In a further aspect, provided herein are methods of reducing neutrophil,inflammatory dendritic cell (iDC), and/or CD4 T cell accumulation in asubject in need thereof comprising administering to the subject aneffective amount of an antagonist of Chemokine Receptor 6 (CCR6) and/orC-X-C motif chemokine receptor 2 (CXCR2).

Antagonists of CCR6 and/or CXCR2

In some embodiments, the CCR6 and/or CXCR2 antagonist has the formula:

wherein

-   -   B is selected from the group consisting of furanyl, thiophenyl,        oxazolyl, phenyl, pyridyl, pyrimidinyl and pyrazinyl, each of        which is optionally substituted with R^(1a), R^(1b), and R²        which are independently selected from the group consisting of        halogen, CN, C₁₋₄ alkyl, C₁₋₄ alkoxy and C₁₋₄ haloalkyl;    -   R³ is a member selected from the group consisting of H and D;    -   R⁴ is a member selected from the group consisting of H, C₁₋₈        alkyl, OH, —NR^(a)R^(b), —C₁₋₄ alkoxy, and Y; wherein the C₁₋₈        alkyl is optionally substituted with halogen, —CN, —CO₂R^(a),        —CONR^(a)R^(b), —C(O)R^(a), OC(O)NR^(a)R^(b), —NR^(a)C(O)R^(b),        —NR^(a)C(O)₂R^(c), —NR^(a)C(O)NR^(a)R^(b), —NR^(a)R^(b),        —OR^(a), —S(O)₂NR^(a)R^(b), —NR^(a)S(O)₂R^(b) and Y, wherein Y        is a 4 to 8 membered cycloheteroalkyl group or a 3 to 8 membered        cycloalkyl group or a 5- or 6-membered aryl or heteroaryl group        any of which is optionally substituted with from 1 to four        substituents selected from halogen, oxo, —CN, —C₁₋₆ alkyl, —C₁₋₆        alkoxy, —C₁₋₆ hydroxyalkyl, —C₁₋₆ haloalkyl, O—C₁₋₆ haloalkyl,        —C₁₋₄alkyl-O—C₁₋₄ alkyl, —C₁₋₆ alkyl-NR^(a)R^(b) , —C₁₋₆        alkyl-CO₂H, —C₁₋₆ alkyl-CO₂R^(a), —C₁₋₆ alkyl-CONR^(a)R^(b),        —C₁₋₆ alkyl-C(O)R^(a), —C₁₋₆ alkyl-OC(O)NR^(a)R^(b), —C₁₋₆        alkyl-NR^(a)C(O)R^(b), —C₁₋₆ alkyl-NR^(a)C(O)₂R^(c), —C₁₋₆        alkyl-NR^(a)C(O)NR^(a)R^(b), —C₁₋₆ alkyl-OR^(a), —C₁₋₆        alkyl-S(O)₂NR^(a)R^(b), —C₁₋₆ alkyl-NR^(a)S(O)₂R^(b) ,        —CO₂R^(a), —CONR^(a)R^(b), —C(O)R^(a), —OC(O)NR^(a)R^(b),        —NR^(a)C(O)R^(b), —NR^(a)C(O)₂R^(c), —NR^(a)C(O)NR^(a)R^(b),        —NR^(a)R^(b), —OR^(a), —S(O)₂NR^(a)R^(b), —NR^(a)S(O)₂R^(b),        —CH₂CO₂R^(a); each R^(a) and R^(b) is independently selected        from hydrogen, C₁₋₄ alkyl, C₁₋₄hydroxyalkyl and C₁₋₄ haloalkyl,        and R^(c) is selected from C₁₋₄ alkyl, C₁₋₄ hydroxyalkyl and        C₁₋₄ haloalkyl; and wherein the 4 to 8 membered cycloheteroalkyl        group and the 3 to 8 membered cycloalkyl group may additionally        be optionally substituted with oxo;    -   R^(5a) and R^(5b) are each members independently selected from        the group consisting of H, halogen, C₁₋₄ alkyl, —C₁₋₄ haloalkyl,        O—C₁₋₄ haloalkyl, C₁₋₄ alkoxy, CO₂H and CN;    -   R^(6a) and R^(6b) are each members independently selected from        the group consisting of H, C₁₋₄ alkyl, C₁₋₄ hydroxyalkyl and        C₁₋₄ haloalkyl; or optionally R^(6a) and R^(6b) are taken        together to form oxo (═O) or a 4 to 6 membered cycloheteroalkyl        group or a 3 to 6 membered cycloalkyl group;    -   R⁷ is a member selected from the group consisting of methyl,        ethyl and C₁₋₂ haloalkyl; and the subscript n is 1 or 2;    -   or any pharmaceutically acceptable salts, solvates, hydrates,        N-oxides, tautomers or rotamers thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist has the formula:

-   -   or any salts, solvates, hydrates, N-oxides, tautomers or        rotamers thereof, wherein    -   B is selected from the group consisting of furanyl, oxazolyl,        phenyl, pyridyl, pyrimidinyl and pyrazinyl, each of which is        optionally substituted with R^(1a), R^(1b), and R² which are        independently selected from the group consisting of halogen, CN,        C₁₋₄ alkyl, C₁₋₄ alkoxy and C₁₋₄ haloalkyl;    -   R³ is a member selected from H and D;    -   R⁴ is a member selected from H, C₁₋₈ alkyl, and Y; wherein the        C₁₋₈ alkyl is optionally substituted with halogen, —CN,        —CO₂R^(a), —CONR^(a)R^(b), —C(O)R^(a), OC(O)NR^(a)R^(b),        —NR^(a)C(O)R^(b), —NR^(a)C(O)₂R^(c), —NR^(a)C(O)NR^(a)R^(b),        —NR^(a)R^(b), —OR^(a), —S(O)₂NR^(a) _(R) ^(b), —NR^(a)S(O)₂R^(b)        and Y, wherein each R^(a) and R^(b) is independently selected        from hydrogen, C₁₋₄ alkyl, C₁₋₄ hydroxyalkyl and C₁₋₄ haloalkyl,        R^(c) is selected from C₁₋₄ alkyl, C₁₋₄ hydroxyalkyl and C₁₋₄        haloalkyl, and Y is a 5 or 6 membered aryl or heteroaryl group        optionally substituted with from one to four substituents        selected from halogen, —CN, —C₁₋₄ alkyl, —C₁₋₄ alkoxy, —C₁₋₄        hydroxyalkyl, —C₁₋₄ haloalkyl, OCF₃, —CO₂R^(a), —CONR^(a)b^(b),        —C(O)R^(a), —OC(O)NR^(a)R^(b), —NR^(a)C(O)R^(b), —CH₂CO₂R^(a);    -   R^(5a) and R^(5b) are each members independently selected from        H, halogen, C₁₋₄ alkyl, C₁₋₄ alkoxy, CO₂H and CN;    -   R^(6a) and R^(6b) are each members independently selected from        H, C₁₋₄ alkyl, C₁₋₄ hydroxyalkyl and C₁₋₄ haloalkyl; or        optionally R^(6a) and R^(6b) are taken together to form oxo        (═O); and the subscript n is 1 or 2.

In some embodiments, the CCR6 and/or CXCR2 antagonist has the formula:

wherein R^(1a) is selected from CH₃ and Cl; R^(1b) is H or is CH₃; R³ isH or D; R⁴ is H or Y; R^(5a) and R^(5b) are each independently selectedfrom H, F, Cl, Br and CH₃; R^(6a) and R^(6b) are each independentlyselected from H and CH₃; and R⁷ is methyl or ethyl; or apharmaceutically acceptable salt, solvate or hydrate, thereof.

In some embodiments, R^(1a) is CH₃; R^(1b) is absent or is CH₃; R³ is Hor D; R⁴ is H; R^(5a) is H, F, Me or Cl or Br; R^(5b) is H or F; R^(6a)and R^(6b) are each H; and R⁷ is methyl or ethyl; or a pharmaceuticallyacceptable salt, solvate or hydrate, thereof.

In some embodiments, the compound is substantially free of other isomersat the carbon atom bearing R³.

In some embodiments, R⁴ is Y.

In some embodiments, a compound of formula (A2) is provided:

wherein R^(1a) is selected from CH₃ and Cl; R^(1b) is H or CH₃; R³ is Hor D; R^(4a) and R^(4b) are independently selected from halogen, —CN,—C₁₋₄ alkyl, —C₁₋₄ alkoxy, —C₁₋₄ hydroxyalkyl, —C₁₋₄ haloalkyl, OCF₃,—CO₂R^(a), —CONR^(a)R^(b), —C(O)R^(a), —OC(O)NR^(a)R^(b),—NR^(a)C(O)R^(b), —CH₂CO₂R^(a), and R^(a) and R^(b) are independentlyselected from hydrogen, C₁₋₄ alkyl, C₁₋₄ hydroxyalkyl and C₁₋₄haloalkyl; R^(5a) and R^(5b) are each independently selected from H, F,Cl, Br and CH₃; R^(6a) and R^(6b) are each independently selected from Hand CH₃; and R⁷ is selected from the group consisting of methyl, ethyland C₁-₂ haloalkyl; or a pharmaceutically acceptable salt, solvate orhydrate, thereof.

In some embodiments, a compound, or a pharmaceutically acceptable saltthereof, is provided, selected from the group consisting of:

In some embodiments, a compound, or a pharmaceutically acceptable saltthereof, is provided, selected from the group consisting of:

In some embodiments, a compound, or a pharmaceutically acceptable saltthereof, is provided, selected from the group consisting of:

In some selected embodiments, compounds of formula (I) are provided thatare selected from those compounds in FIG. 1 .

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.129:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.123:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.136:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is compound 1.138:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the CCR6 and/or CXCR2 antagonist is selected fromthe compounds or pharmaceutical compositions disclosed in U.S. Pat. No.9,834,545, stemming from application Ser. No. 15/353,889, filed on Nov.17, 2016 by ChemoCentryx, the content of which is incorporated hereinfor all purposes.

Pharmaceutical Compositions

In addition the compounds described above, the compositions formodulating CCR6 and/or CXCR2 activity in humans and animals willtypically contain a pharmaceutical carrier or diluent.

The term “composition” as used herein is intended to encompass a productcomprising the specified ingredients in the specified amounts, as wellas any product which results, directly or indirectly, from combinationof the specified ingredients in the specified amounts. By“pharmaceutically acceptable” it is meant the carrier, diluent orexcipient must be compatible with the other ingredients of theformulation and not deleterious to the recipient thereof.

The pharmaceutical compositions for the administration of the compoundsof this invention may conveniently be presented in unit dosage form andmay be prepared by any of the methods well known in the art of pharmacyand drug delivery. All methods include the step of bringing the activeingredient into association with the carrier which constitutes one ormore accessory ingredients. In general, the pharmaceutical compositionsare prepared by uniformly and intimately bringing the active ingredientinto association with a liquid carrier or a finely divided solid carrieror both, and then, if necessary, shaping the product into the desiredformulation. In the pharmaceutical composition the active objectcompound is included in an amount sufficient to produce the desiredeffect upon the process or condition of diseases.

The pharmaceutical compositions containing the active ingredient may bein a form suitable for oral use, for example, as tablets, troches,lozenges, aqueous or oily suspensions, dispersible powders or granules,emulsions and self emulsifications as described in U.S. Pat. No.6,451,339, hard or soft capsules, syrups, elixirs, solutions, buccalpatch, oral gel, chewing gum, chewable tablets, effervescent powder andeffervescent tablets. Compositions intended for oral use may be preparedaccording to any method known to the art for the manufacture ofpharmaceutical compositions and such compositions may contain one ormore agents selected from the group consisting of sweetening agents,flavoring agents, coloring agents, antioxidants and preserving agents inorder to provide pharmaceutically elegant and palatable preparations.Tablets contain the active ingredient in admixture with non-toxicpharmaceutically acceptable excipients which are suitable for themanufacture of tablets. These excipients may be for example, inertdiluents, such as cellulose, silicon dioxide, aluminum oxide, calciumcarbonate, sodium carbonate, glucose, mannitol, sorbitol, lactose,calcium phosphate or sodium phosphate; granulating and disintegratingagents, for example, corn starch, or alginic acid; binding agents, forexample PVP, cellulose, PEG, starch, gelatin or acacia, and lubricatingagents, for example magnesium stearate, stearic acid or talc. Thetablets may be uncoated or they may be coated, enterically or otherwise,by known techniques to delay disintegration and absorption in thegastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate may be employed. They may also becoated by the techniques described in the U.S. Pat. Nos. 4,256,108;4,166,452; and 4,265,874 to form osmotic therapeutic tablets for controlrelease.

Formulations for oral use may also be presented as hard gelatin capsuleswherein the active ingredient is mixed with an inert solid diluent, forexample, calcium carbonate, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, for example peanut oil, liquid paraffin, or olive oil.Additionally, emulsions can be prepared with a non-water miscibleingredient such as oils and stabilized with surfactants such asmono-diglycerides, PEG esters and the like.

Aqueous suspensions contain the active materials in admixture withexcipients suitable for the manufacture of aqueous suspensions. Suchexcipients are suspending agents, for example sodiumcarboxymethylcellulose, methylcellulose, hydroxy-propylmethylcellulose,sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia;dispersing or wetting agents may be a naturally-occurring phosphatide,for example lecithin, or condensation products of an alkylene oxide withfatty acids, for example polyoxy-ethylene stearate, or condensationproducts of ethylene oxide with long chain aliphatic alcohols, forexample heptadecaethyleneoxycetanol, or condensation products ofethylene oxide with partial esters derived from fatty acids and ahexitol such as polyoxyethylene sorbitol monooleate, or condensationproducts of ethylene oxide with partial esters derived from fatty acidsand hexitol anhydrides, for example polyethylene sorbitan monooleate.The aqueous suspensions may also contain one or more preservatives, forexample ethyl, or n-propyl, p-hydroxybenzoate, one or more coloringagents, one or more flavoring agents, and one or more sweetening agents,such as sucrose or saccharin.

Oily suspensions may be formulated by suspending the active ingredientin a vegetable oil, for example arachis oil, olive oil, sesame oil orcoconut oil, or in a mineral oil such as liquid paraffin. The oilysuspensions may contain a thickening agent, for example beeswax, hardparaffin or cetyl alcohol. Sweetening agents such as those set forthabove, and flavoring agents may be added to provide a palatable oralpreparation. These compositions may be preserved by the addition of ananti-oxidant such as ascorbic acid.

Dispersible powders and granules suitable for preparation of an aqueoussuspension by the addition of water provide the active ingredient inadmixture with a dispersing or wetting agent, suspending agent and oneor more preservatives. Suitable dispersing or wetting agents andsuspending agents are exemplified by those already mentioned above.Additional excipients, for example sweetening, flavoring and coloringagents, may also be present.

The pharmaceutical compositions of the invention may also be in the formof oil-in-water emulsions. The oily phase may be a vegetable oil, forexample olive oil or arachis oil, or a mineral oil, for example liquidparaffin or mixtures of these. Suitable emulsifying agents may benaturally-occurring gums, for example gum acacia or gum tragacanth,naturally-occurring phosphatides, for example soy bean, lecithin, andesters or partial esters derived from fatty acids and hexitolanhydrides, for example sorbitan monooleate, and condensation productsof the said partial esters with ethylene oxide, for examplepolyoxyethylene sorbitan monooleate. The emulsions may also containsweetening and flavoring agents.

Syrups and elixirs may be formulated with sweetening agents, for exampleglycerol, propylene glycol, sorbitol or sucrose. Such formulations mayalso contain a demulcent, a preservative, flavoring and/or coloringagents. Oral solutions can be prepared in combination with, for example,cyclodextrin, PEG and surfactants.

The pharmaceutical compositions may be in the form of a sterileinjectable aqueous or oleagenous suspension. This suspension may beformulated according to the known art using those suitable dispersing orwetting agents and suspending agents which have been mentioned above.The sterile injectable preparation may also be a sterile injectablesolution or suspension in a non-toxic parenterally-acceptable diluent orsolvent, for example as a solution in 1,3-butane diol. Among theacceptable vehicles and solvents that may be employed are water,Ringer's solution and isotonic sodium chloride solution. In addition,sterile, fixed oils are conventionally employed as a solvent orsuspending medium. For this purpose any bland fixed oil may be employedincluding synthetic mono- or diglycerides. In addition, fatty acids suchas oleic acid find use in the preparation of injectables.

The compounds of the present invention may also be administered in theform of suppositories for rectal administration of the drug. Thesecompositions can be prepared by mixing the drug with a suitablenon-irritating excipient which is solid at ordinary temperatures butliquid at the rectal temperature and will therefore melt in the rectumto release the drug. Such materials include cocoa butter andpolyethylene glycols. Additionally, the compounds can be administeredvia ocular delivery by means of solutions or ointments. Still further,transdermal delivery of the subject compounds can be accomplished bymeans of iontophoretic patches and the like. For topical use, creams,ointments, jellies, solutions or suspensions, etc., containing thecompounds of the present invention are employed. As used herein, topicalapplication is also meant to include the use of mouth washes andgargles.

The compounds of the invention may be formulated for depositing into amedical device, which may include any of variety of conventional grafts,stents, including stent grafts, catheters, balloons, baskets or otherdevice that can be deployed or permanently implanted within a bodylumen. As a particular example, it would be desirable to have devicesand methods which can deliver compounds of the invention to the regionof a body which has been treated by interventional technique.

In exemplary embodiment, the inhibitory agent of this invention may bedeposited within a medical device, such as a stent, and delivered to thetreatment site for treatment of a portion of the body.

Stents have been used as delivery vehicles for therapeutic agents (i.e.,drugs). Intravascular stents are generally permanently implanted incoronary or peripheral vessels. Stent designs include those of U.S. Pat.No. 4,733,655 (Palmaz), U.S. Pat. No. 4,800,882 (Gianturco), or U.S.Pat. No. 4,886,062 (Wiktor). Such designs include both metal andpolymeric stents, as well as self-expanding and balloon-expandablestents. Stents may also used to deliver a drug at the site of contactwith the vasculature, as disclosed in U.S. Pat. No. 5,102,417 (Palmaz)and in International Patent Application Nos. WO 91/12779 (Medtronic,Inc.) and WO 90/13332 (Cedars-Sanai Medical Center), U.S. Pat. Nos.5,419,760 (Narciso, Jr.) and U.S. Pat. No. 5,429,634 (Narciso, Jr.), forexample. Stents have also been used to deliver viruses to the wall of alumen for gene delivery, as disclosed in U.S. Pat. No. 5,833,651(Donovan et al.).

In one embodiment, the inhibitory agent may be incorporated with polymercompositions during the formation of biocompatible coatings for medicaldevices, such as stents. The coatings produced from these components aretypically homogeneous and are useful for coating a number of devicesdesigned for implantation.

The polymer may be either a biostable or a bioabsorbable polymerdepending on the desired rate of release or the desired degree ofpolymer stability, but a bioabsorbable polymer is preferred for thisembodiment since, unlike a biostable polymer, it will not be presentlong after implantation to cause any adverse, chronic local response.Bioabsorbable polymers that could be used include, but are not limitedto, poly(L-lactic acid), polycaprolactone, polyglycolide (PGA),poly(lactide-co-glycolide) (PLLA/PGA), poly(hydroxybutyrate),poly(hydroxybutyrate-co-valerate), polydioxanone, polyorthoester,polyanhydride, poly(glycolic acid), poly(D-lactic acid), poly(L-lacticacid), poly(D,L-lactic acid), poly(D,L-lactide) (PLA) , poly (L-lactide)(PLLA), poly(glycolic acid-co-trimethylene carbonate) (PGA/PTMC),polyethylene oxide (PEO), polydioxanone (PDS), polyphosphoester,polyphosphoester urethane, poly(amino acids), cyanoacrylates,poly(trimethylene carbonate), poly(iminocarbonate), copoly(ether-esters)(e.g., PEO/PLA), polyalkylene oxalates, polyphosphazenes andbiomolecules such as fibrin, fibrinogen, cellulose, starch, collagen andhyaluronic acid, polyepsilon caprolactone, polyhydroxy butyric acid,polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates,cross linked or amphipathic block copolymers of hydrogels, and othersuitable bioabsorbable poplymers known in the art. Also, biostablepolymers with a relatively low chronic tissue response such aspolyurethanes, silicones, and polyesters could be used and otherpolymers could also be used if they can be dissolved and cured orpolymerized on the medical device such as polyolefins, polyisobutyleneand ethylene-alphaolefin copolymers; acrylic polymers and copolymers,vinyl halide polymers and copolymers, such as polyvinyl chloride;polyvinylpyrrolidone; polyvinyl ethers, such as polyvinyl methyl ether;polyvinylidene halides, such as polyvinylidene fluoride andpolyvinylidene chloride; polyacrylonitrile, polyvinyl ketones; polyvinylaromatics, such as polystyrene, polyvinyl esters, such as polyvinylacetate; copolymers of vinyl monomers with each other and olefins, suchas ethylene-methyl methacrylate copolymers, acrylonitrile-styrenecopolymers, ABS resins, and ethylene-vinyl acetate copolymers; pyrancopolymer; polyhydroxy-propyl-methacrylamide-phenol;polyhydroxyethyl-aspartamide-phenol; polyethyleneoxide-polylysinesubstituted with palmitoyl residues; polyamides, such as Nylon 66 andpolycaprolactam; alkyd resins, polycarbonates; polyoxymethylenes;polyimides; polyethers; epoxy resins, polyurethanes; rayon;rayon-triacetate; cellulose, cellulose acetate, cellulose butyrate;cellulose acetate butyrate; cellophane; cellulose nitrate; cellulosepropionate; cellulose ethers; and carboxymethyl cellulose.

Polymers and semipermeable polymer matrices may be formed into shapedarticles, such as valves, stents, tubing, prostheses and the like.

In one embodiment of the invention, the inhibitory agent of theinvention is coupled to a polymer or semipermeable polymer matrix thatis formed as a stent or stent-graft device.

Typically, polymers are applied to the surface of an implantable deviceby spin coating, dipping or spraying. Additional methods known in theart can also be utilized for this purpose.

Methods of spraying include traditional methods as well asmicrodeposition techniques with an inkjet type of dispenser.Additionally, a polymer can be deposited on an implantable device usingphoto-patterning to place the polymer on only specific portions of thedevice. This coating of the device provides a uniform layer around thedevice which allows for improved diffusion of various analytes throughthe device coating.

In preferred embodiments of the invention, the inhibitory agent isformulated for release from the polymer coating into the environment inwhich the medical device is placed. Preferably, the inhibitory agent isreleased in a controlled manner over an extended time frame (e.g.,months) using at least one of several well-known techniques involvingpolymer carriers or layers to control elution. Some of these techniqueswere previously described in U.S. Patent Application 20040243225A1.

Moreover, as described for example in U.S. Pat. No. 6,770,729, thereagents and reaction conditions of the polymer compositions can bemanipulated so that the release of the inhibitory agent from the polymercoating can be controlled. For example, the diffusion coefficient of theone or more polymer coatings can be modulated to control the release ofthe inhibitory agent from the polymer coating. In a variation on thistheme, the diffusion coefficient of the one or more polymer coatings canbe controlled to modulate the ability of an analyte that is present inthe environment in which the medical device is placed (e.g. an analytethat facilitates the breakdown or hydrolysis of some portion of thepolymer) to access one or more components within the polymer composition(and for example, thereby modulate the release of the inhibitory agentfrom the polymer coating). Yet another embodiment of the inventionincludes a device having a plurality of polymer coatings, each having aplurality of diffusion coefficients. In such embodiments of theinvention, the release of the inhibitory agent from the polymer coatingcan be modulated by the plurality of polymer coatings.

In yet another embodiment of the invention, the release of theinhibitory agent from the polymer coating is controlled by modulatingone or more of the properties of the polymer composition, such as thepresence of one or more endogenous or exogenous compounds, oralternatively, the pH of the polymer composition. For example, certainpolymer compositions can be designed to release a inhibitory agent inresponse to a decrease in the pH of the polymer composition.Alternatively, certain polymer compositions can be designed to releasethe inhibitory agent in response to the presence of hydrogen peroxide.

In some embodiments, a pharmaceutical composition comprising a compoundof the present disclosure is provided. In some embodiments, thepharmaceutical composition further comprises one or more additionaltherapeutic agents. In some embodiments, the one or more additionaltherapeutic agent is selected from the group consisting of cytotoxicchemotherapy, anti-cancer or anti-tumor vaccines, anti-immunocytokinetherapies, immunocytokine therapies, chimeric antigen receptor (CAR) Tcell receptors, gene transfer therapy, checkpoint inhibitors,corticosteroids, retinoid-like agents, antineoplastics, and interferonsanalogs. In some embodiments, the one or more additional therapeuticagent is selected from the group consisting of a TNF alpha ligandinhibitor, a TNF binding agent, an IL-1 ligand inhibitor; an IL-6 ligandinhibitor, an IL-8 ligand inhibitor; an IL-17 antagonist, a calcineurininhibitor, a TNF antagonist, a Retinoic acid receptor gamma antagonist,an IL-17A ligand inhibitor; an IL-17F ligand inhibitor, a RIP-1 kinaseinhibitor, a sphingosine-1-phosphate receptor-1 antagonist, asphingosine-1-phosphate receptor-1 modulator, a Rho associated proteinkinase 2 inhibitor, an IL-12 antagonist; an IL-23 antagonist, a type IITNF receptor modulator, an IL-23A inhibitor, a PDE 4 inhibitor, a JAKtyrosine kinase inhibitor, a Jakl tyrosine kinase inhibitor; a Jak3tyrosine kinase inhibitor, a Histamine H1 receptor antagonist, aRetinoic acid receptor agonist, a membrane copper amine oxidaseinhibitor, a PI3K modulator, a Phosphoinositide-3 kinase deltainhibitor, a mitochondrial 10 kDa heat shock protein stimulator, anadenosine A3 receptor agonist, a galectin-3 inhibitor, a FIFO ATPsynthase modulator, a GM-CSF ligand inhibitor, a vitamin D3 receptoragonist, a glucocorticoid agonist, a histamine H4 receptor antagonist, aCCR3 chemokine antagonist, an eotaxin ligand inhibitor, aSphingosine-1-phosphate receptor-1 modulator, a phospholipase A2inhibitor, a PDE 4 inhibitor, an albumin modulator, a TLR-7 antagonist,a TLR-8 antagonist a TLR-9 antagonist, a CD40 ligand receptorantagonist, a Src tyrosine kinase inhibitor, a tubulin binding agent, aninterleukin-1 alpha ligand inhibitor, a histone deacetylase-1 inhibitor,a histone deacetylase-2 inhibitor, a histone deacetylase-3 inhibitor, ahistone deacetylase-6 inhibitor, a nucleoside reverse transcriptaseinhibitor, a nuclear factor kappa B inhibitor, a STAT-3 inhibitor, aparathyroid hormone ligand inhibitor; a vitamin D3 receptor agonist, a Tcell surface glycoprotein CD28 stimulator, a histamine H4 receptorantagonist, a TGF beta agonist, a P-selectin glycoprotein ligand-1stimulator, a DHFR inhibitor, a Retinoic acid receptor gamma modulator,a cytosolic phospholipase A2 inhibitor, a retinoid X receptor modulator,a beta-catenin inhibitor, a CREB binding protein inhibitor, a TrkAreceptor antagonist, a T-cell differentiation antigen CD6 inhibitor, anADP ribosyl cyclase-1 inhibitor, an Interleukin-1 beta ligand modulator;an insulin receptor substrate-1 inhibitor, a DHFR inhibitor, an IL-8antagonist, a drug that blocks the activity of CTLA-4 (CD152), PD-1(CD279), PDL-1 (CD274), TIM-3, LAG-3 (CD223), VISTA, KIR, NKG2A, BTLA,PD-1H, TIGIT, CD96, 4-1BB (CD137), 4-1BBL (CD137L), GARP, CSF-1R, A2AR,CD73, CD47, tryptophan 2,3-dioxygenase (TDO) or indoleamine 2,3dioxygenase (IDO), and agonists of OX40, GITR, 4-1BB, ICOS, STING orCD40.

Methods of Administration

In general, treatment methods provided herein comprise administering toa patient an effective amount of one or more compounds provided herein.In a preferred embodiment, the compound(s) of the invention arepreferably administered to a patient (e.g., a human) orally ortopically. Treatment regimens may vary depending on the compound used Ingeneral, a dosage regimen of 2 times daily is more preferred, with oncea day dosing particularly preferred. It will be understood, however,that the specific dose level and treatment regimen for any particularpatient will depend upon a variety of factors including the activity ofthe specific compound employed, the age, body weight, general health,sex, diet, time of administration, route of administration, rate ofexcretion, drug combination (i.e., other drugs being administered to thepatient) and the severity of the particular disease undergoing therapy,as well as the judgment of the prescribing medical practitioner. Ingeneral, the use of the minimum dose sufficient to provide effectivetherapy is preferred. Patients may generally be monitored fortherapeutic effectiveness using medical or veterinary criteria suitablefor the condition being treated or prevented.

The compounds and compositions of the present invention may beadministered by oral, parenteral (e.g., intramuscular, intraperitoneal,intravenous, ICV, intracisternal injection or infusion, subcutaneousinjection, or implant), inhalation, nasal, vaginal, rectal, sublingual,or topical routes of administration and may be formulated, alone ortogether, in suitable dosage unit formulations containing conventionalnontoxic pharmaceutically acceptable carriers, adjuvants and vehiclesappropriate for each route of administration. The present invention alsocontemplates administration of the compounds and compositions of thepresent invention in a depot formulation.

Dosage levels of the order of from about 0.1 mg to about 100 mg perkilogram of body weight per day are useful in the treatment orpreventions of conditions involving pathogenic CCR6 and/or CXCR2activity (about 0.5 mg to about 7 g per human patient per day).Preferably, the dosage level will be about 0.01 to about 25 mg/kg perday; more preferably about 0.05 to about 10 mg/kg per day. A suitabledosage level may be about 0.01 to 25 mg/kg per day, about 0.05 to 10mg/kg per day, or about 0.1 to 5 mg/kg per day. Within this range thedosage may be 0.005 to 0.05, 0.05 to 0.5, 0.5 to 5.0, or 5.0 to 50 mg/kgper day. For oral administration, the compositions are preferablyprovided in the form of tablets containing 1.0 to 1000 milligrams of theactive ingredient, particularly 1.0, 5.0, 10.0, 15.0, 20.0, 25.0, 50.0,75.0, 100.0, 150.0, 200.0, 250.0, 300.0, 400.0, 500.0, 600.0, 750.0,800.0, 900.0, and 1000.0 milligrams of the active ingredient for thesymptomatic adjustment of the dosage to the patient to be treated. Thecompounds may be administered on a regimen of 1 to 4 times per day,preferably once or twice per day.

The amount of active ingredient that may be combined with the carriermaterials to produce a single dosage form will vary depending upon thehost treated and the particular mode of administration. Dosage unitforms will generally contain between from about 1 mg to about 500 mg ofan active ingredient. For compounds administered orally, transdermally,intravaneously, or subcutaneously, it is preferred that sufficientamount of the compound be administered to achieve a serum concentrationof 5 ng (nanograms)/mL-10 μg (micrograms)/mL serum, more preferablysufficient compound to achieve a serum concentration of 20 ng-1 μg/mlserum should be administered, most preferably sufficient compound toachieve a serum concentration of 50 ng/ml-200 ng/ml serum should beadministered.

It will be understood, however, that the specific dose level andfrequency of dosage for any particular patient may be varied and willdepend upon a variety of factors including the activity of the specificcompound employed, the metabolic stability and length of action of thatcompound, the age, body weight, hereditary characteristics, generalhealth, sex, diet, mode and time of administration, rate of excretion,drug combination, the severity of the particular condition, and the hostundergoing therapy.

I. EXAMPLES Example 1: IL36-Mediated Skin Inflammation RequiresSignaling Through Chemokine Receptor CCR6

Generalized pustular psoriasis (GPP) is a rare inflammatory skindisorder with an etiology distinct from the more common plaquepsoriasis. GPP patients often do not respond to therapeutic agentstypically used for plaque psoriasis. Genetic evidence suggests that GPParises from dysfunctions in the IL36/IL36Ra/IL36R signaling axis, andmany aspects of GPP can be re-created through intradermal (ID) injectionof pre-activated IL36 in the mouse. We have used this ID-IL36 model tostudy the leukocyte populations accumulating within GPP skin. In aprevious study, we reported that small molecule CCR6 antagonist Compound1.129 ameliorates inflammation in a topical imiquimod-induced model ofplaque psoriasis. CCR6 antagonism prevents accumulation ofIL17-secreting γδT (γδT17) in this model, which appears to be themechanism by which Compound 1.129 alleviates disease. We have found inthe present study that γδT17 cells do not accumulate in IL36-inflamedskin. Instead, a conventional CD4⁺αβ T cell population accumulates in inthis model. Compound 1.129 reduces IL36-induced inflammation whilepreventing accumulation of this CD4⁺αβ T cell population. Thus, althoughdisparate T cell populations are associated with each model,inflammation is ameliorated in both by CCR6 antagonism and CXCR2antagonism. These findings suggest that CCR6 may constitute a noveltarget for a mechanistically distinct approach for GPP therapy.

Example 2: Inhibition of Chemokine receptor and Ligand InteractionsReverses IL-36-Induced Inflammation

Materials and Methods

For in vitro determination of chemokine receptor inhibitor activity,Compound 1.136 was dissolved in DMSO to generate a stock concentrationof 10 mM that was further diluted in chemotaxis migration buffer (HBSS,1% BSA, 1% HEPES) to create a 10-point inhibitor gradient ranging from100 nM-0.01 nM. Migration assay was then run in 100% mouse serum inresponse to rmCCL20 for CCR6 activity or rmCXCL1 for CXCR2 activityusing ChemoTx plates (Neuroprobe, Gaithersburg Md.) and assessed byfluorescence staining of migrated cells with CyQuant (Thermo Fisher).

Balb/c mice were purchased from Jackson Laboratories (Bar Harbor, Maine)and housed at ChemoCentryx animal facility in accordance with guidelinesdescribed in the Guide and Use of Laboratory Animals of the NationalResearch Council. All studies were approved by the ChemoCentryxInstitutional Animal Care and Use Committee.

For in vivo dosing, mice were dosed subcutaneously with Compound 1.136once daily (s.c. q.d. in 1% HPMC) at 90 mg/kg unless stated otherwise,starting at day 0 of a study. The in vivo dose was predicted based on invitro potency and the pharmacokinetic response to a single dose in mice.The minimum antagonist concentration (at trough) to fully cover agradient-sensing chemoattractant receptor has been determined tocorrespond with the IC₉₀ concentration (Schall, T. J., and A. E.Proudfoot. 2011. Overcoming hurdles in developing successful drugstargeting chemokine receptors. Nat Rev Immunol 11: 355-363). The IC₉₀ ofCompound 1.136 in 100% serum is 45 ng/ml for CCR6 inhibition and 145ng/ml for CXCR2 inhibition. The actual trough concentrations achieved invivo for Compound 1.136 at 30, 60 and 90 mg/kg are shown in Table 1.

TABLE 1 Mean ± s.d. plasma concentrations measured at trough (i.e. 24hours post final dose (day 5)) for animals dosed with 90, 60, 30 mg/kgCompound 1.136 in 1% HPM Compound 1.136 Dose (mg/kg)  90 mg/kg  60 mg/kg 30 mg/kg Compound 1.136 973 ± 19 323 ± 20 267 ± 18 Trough Value (ng/ml)

Mouse model of IL-36-induced ear thickening. The right ear of 8-week oldBalb/c mice received intradermal injections of recombinant murine IL-36α(150 μg per mouse formulated in PBS; BioLegend) every day for 5 days.Intradermal injections of PBS were given to the left ear as a control.Ear thickness was measured prior to start of study and each daythoughout the study using a digital micrometer. The imiquimod model ofpsoriasiform dermatitis was performed as previously described (CampbellJet al. J Immunol. 2017; 199(9):3129-36. van der Fits et al. J Immunol2009; 182(9):5836-45). Briefly, 5% Imiquimod (Fougera®) was applieddaily to the shaved and depilated backs of Balb/c mice for up to 4 days.Control mice were treated with an application of Vaseline. Erythema,desquamation, and skin thickening were scored independently on a scalefrom 0 to 4 where 0=no disease; 1=slight disease; 2=moderate disease;3=marked disease; 4=very marked disease (Campbell J et al. J Immunol.2017; 199(9):3129-36. van der Fits et al. J Immunol 2009;182(9):5836-45). Skin from the backs of mice was excised at the end ofthe study and processed for flow cytometry analysis,

Flow cytometry of leukocytes. Ears or excised skin samples were digestedin collagenase A and 1 U/ml DNAse I with agitation for 30 min at 37° C.Cells were then dislodged from skin, filtered through a 70 μM sieve,washed and re-suspended in FACS buffer (PBS with 10% FBS) for analysis.

Directly conjugated Monoclonal antibodies were from R&D Systems(Minneapolis, Minn.), BioLegend (San Diego, Calif.), oreBioscience/ThermoFisher (San Diego, Calif.). CD45.2 (104) inAlexaFluor488, Ly6C (HK1.4) in PE, CD90.2 (30-H12) in Per CP-Cy5.5,TCRVγ4 (UC3-10A6) in PE-Cy7, TCRβ (H57-597) in APC, CD4 (GK1.5) inAPC-eFluor-780, TCRγδ (GL3) in BV-421, LIVE/DEAD fixable Aqua stain(Molecular Probes, Eugene, Oreg.), CD11c (N418) in BV650, Ly6G (1A4) inBV711 and CD11b (M1/70) in BV785. Flow cytometry data were acquired witha Fortessa (BD Biosciences) cytometer and analyzed using FlowJo v10.2

Flow cytometry of leukocytes extracted from inflamed skin shown in FIG.5 and FIG. 11 involved a panel of unconjugated monoclonals (or theirisotype-matched controls) followed by an appropriate APC-conjugatedpolyclonal (FAB)₂ (Jackson ImmunoResearch, West Grove, Pa., USA). Theseunconjugated reagents included anti-CCR6 (140706) from R&D Systems(Minneapolis, Minn.), and anti-CXCR2 (SA044G4) from Biolegend (SanDiego, Calif.) all rat IgG2as. Unconjugated MAbs against lineage markersCD103, CD11c, FLT3, CD205, and F4/80 were all obtained from Biolegend.After staining with unconjugated monoclonal and APC polyclonal, thecells were blocked with 10% mouse, 10% hamster and 10% rat sera (JacksonImmunoResearch) prior to staining with direct conjugates.

Mouse anti-IL-17RA. (R&D Systems) was dosed as a positive control at200-500 μg per mouse, i.p., q.d. The 200 μg/ml per mouse per day dosingwas found to be well within the plateau of maximal activity for thistreatment (FIG. 6A).

Multiplex analysis of chemokine protein concentration in skin. Eartissue collected after 4 days of intradermal IL-36α treatments washomogenized in cold PBS containing protease inhibitors (Roche) thencentrifuged to remove debris. The soluble fraction was assayed using amultiplex assay kit according to manufacturers instructions (R&Dsystems) and read on a MagPix (Luminex) analyzer. The concentration oftissue chemokines was normalized against the total protein levelsmeasured for each sample using the standard Bradford assay.

For hematoxylin and eosin (H&E) staining, ear tissue was initially fixedin paraformaldehyde for paraffin embedding. Samples were processed bystandard procedures. Epidermal thickness was measured as the average of7 measurements made along the center third (the area containing thelesion) of the length of the H&E stained ear sections using PhotoshopCS4 software (Adobe®)

Statistical significance was determined by Mann Whitney calculationusing GraphPad Prism 6.0 software.

Results & Discussion

Inflammatory cell populations were compared within inflamed skinobtained from two murine models of psoriasis. For plaque psoriasis, thewell-established imiquimod (IMQ) model was used (van der Fits et al. JImmunol 2009; 182(9):5836-45), in which the TLR 7/8 agonist IMQ isapplied daily to the surface of depilated skin. For GPP, dailyintra-cutaneous injections of activated mouse IL-36α to the ear wasused.

After 4 days of treatment flow cytometry showed inflammatory cellinfiltrates isolated from skin (Gated as shown in FIG. 2 ) differappreciably between the two models. IMQ treatment generated a largeneutrophil population (FIG. 3A & FIG. 3C) whereas activated IL-36αgenerated nearly equal numbers of T cells, neutrophils andLy6C^(hi)/Ly6G⁻ myeloid cells (FIG. 3B & FIG. 3C). The T cellpopulations from each model were also highly divergent FIG. 3D);IMQ-treated skin (grey bars with horizontal stripes) having a prominentγδT17 (Vγ4⁺) population (as previously observed (Cai et al. Immunity2011; 35(4):596-610. Campbell J et al. J Immunol. 2017;199(9):3129-36.)), and T cells from IL-36α-injected skin (grey bars withdiagonal stripes) consisted almost entirely of CD4⁺ “conventional” αβ Tcells.

The myeloid skin population induced by IL-36 expressed a combination ofmarkers characteristic of both myeloid and dendritic cells (FIG. 11 ).These cells expressed CD103, CD11c and F4/80. They did not express Flt3,a marker diagnostic of classical dendritic cells (cDC), nor did theyexpress CD205 (DEC205) (FIG. 11 ). Based on this immunophenotype, andthe location of these cells within actively inflamed skin, we believethese cells to be monocyte-derived inflammatory dendritic cells (iDC)(Merad et al. Annu Rev Immunol 2013; 31:563-604.) and will refer to themas such for this point on.

We observed that chemokine ligands for CCL20 and CXCR2 weresignificantly increased in skin by treatment with IL-36α (FIG. 4 ), inagreement with previous work. (Campbell, J. J., K. Ebsworth, L. S. Ertl,J. P. McMahon, D. Newland, Y. Wang, S. Liu, Z. Miao, T. Dang, P. Zhang,I. F. Charo, R. Singh, and T. J. Schall. 2017. IL-17-Secretinggammadelta T Cells Are Completely Dependent upon CCR6 for Homing toInflamed Skin. J Immunol 199: 3129-3136 & van der Fits, L., S. Mourits,J. S. Voerman, M. Kant, L. Boon, J. D. Laman, F. Cornelissen, A. M. Mus,E. Florencia, E. P. Prens, and E. Lubberts. 2009. Imiquimod-inducedpsoriasis-like skin inflammation in mice is mediated via the IL-23/IL-17axis. J Immunol 182: 5836-5845.) We used flow cytometry to examine CCR6and CXCR2 expression on each of the three leukocyte subtypes comprisingmost of the CD45+ infiltrate that accumulated in response to IL-36 (FIG.5 ). Of these three, only neutrophils expressed CXCR2 levels greaterthan isotype control staining (FIG. 5B, left column). A minority of theiDC and a majority of CD4 T cells expressed CCR6 (FIG. 5A, rightcolumn).

The prominence of CCR6 and CXCR2 ligand in the inflamed skin coupledwith the expression of these two receptors by the IL-36α-induced subsetsprompted us to assess a CCR6/CXCR2 antagonist for efficacy in preventingthese cells from accumulating within the skin (and thereby amelioratingIL-36α-induced skin inflammation). Compound 1.136 is a small moleculewith a molecular weight of ˜440 and an IC₅₀ of 10 nM on mouse CCR6 and20 nM on mouse CXCR2 as assessed by in vitro inhibition of chemotaxis in100% serum. The table below provides further information on Compound1.136 in vitro inhibition.

TABLE 2 In vitro Activity of Compound 1.136 CC and CXC ChemokineReceptors Compound mCCR6 ^(a) mCXCR2 ^(b) mCCR1 ^(c) mCCR2 ^(d) mCCR4^(e) mCCR5 ^(f) mCCR7 ^(g) mCCR9 ^(h) mCXCR4 ^(i) 1.136 1020 >10,000 >10,000 >10,000 >10,000 6,000 >10,000 >10,000 ^(a) IC₅₀ forchemotaxis of mCCR6-transfected BaF3 cells to mCCL20 in serum. ^(b) IC₅₀for chemotaxis of mouse bone marrow cells to mCXCL1 in plasma. ^(c) IC₅₀for chemotaxis of mouse WEHI-274.1 cells to mCCL3 in serum. ^(d) IC₅₀for chemotaxis of mouse bone marrow cells to mCCL2 in serum. ^(e) A₂ forchemotaxis of mCCR4-transfected BaF3 cells to mCCL22 in serum. ^(f) IC₅₀for chemotaxis of mCCR5-transfected BaF3 cells to mCCL5 in serum. ^(g)IC₅₀ for chemotaxis of mouse splenocytes to mCCL19 in serum. ^(h) IC₅₀for chemotaxis of mouse thymocytes to mCCL25 in serum. ^(i) IC₅₀ forchemotaxis of mouse BaF3-WT to mCXCL12 in serum.

We measured ear thickness after 4 daily intradermal injections of PBSalone (negative control) or activated IL-36α in PBS (FIG. 6 ). The meanthickness of PBS-treated ears was ˜2.5 mm, half as thick as theIL-36α-treated ears (˜5.0 mm, FIG. 6A). Compound 1.136 wassubcutaneously dosed once daily (on the back of the mouse, distal fromthe IL-36-treated ear), and achieved dose-dependent decreases inIL-36-induced ear thickening (FIG. 6A). A time course revealed that theeffects of Compound 1.136 were appreciable after the second day oftreatment (FIG. 6B and FIG. 7A).

We next compared the effectiveness of Compound 1.136 to that of anα-IL-17RA monoclonal antibody (200 or 500 μg per mouse dosed IP oncedaily). Anti-IL-17RA treatment was effective at reducing IL-36-inducedear swelling (FIG. 6A), but was significantly less effective than the 90mg/kg dose of Compound 1.136. The separation in effectiveness between asaturating dose of anti-IL-17RA MAb and 90 mg/kg Compound 1.136 becameevident after the second day of treatment (FIG. 6B (also, see FIG. 7Bfor direct comparison of anti-IL-17RA MAb treatment to theisotype-matched control for this MAb at 500 μg/mouse/day)).

We next examined sections of mouse ears taken after 4 daily treatmentswith PBS (control) or activated IL-36α for measurement of epidermalthickness (FIG. 8A, FIG. 8B, & FIG. 9 ). In agreement with the directmeasurements of ear swelling shown in FIG. 6 , epidermal thickening wasevident in the IL-36α-treated ear versus the PBS-treated ear (FIG. 9A).Administration of Compound 1.136 or anti-IL-17RA significantly reducedIL-36α-mediated epidermal thickness (FIG. 9B).

Administration of Compound 1.136 reduced IL-36α-induced dermal,epidermal and overall skin thickness (FIG. 8C). In addition, the stratumcorneum remained intact and leukocyte infiltration was reduced. Dosingwith anti-IL-17RA had a beneficial effect on IL-36-treated skin as well(FIG. 8D).

We next assessed the effects of Compound 1.136 on the inflammatory cellsubsets accumulating within IL-36 treated skin by flow cytometry (FIG.10 ). Immune cells were isolated from the skin after 4 daily earinjections of IL-36α and treatment with Compound 1.136 or α-IL-17RA MAb.We found that Compound 1.136 significantly reduced the accumulation ofCD4⁺ T cells, neutrophils and inflammatory iDCs. In contrast, α-IL-17RAdid not reduce the accumulation of CD4⁺ T cells, but had effects similarto Compound 1.136 on neutrophils and iDCs.

Pustular psoriasis is a rare skin disorder comprising several subtypes,including both generalized and localized forms (Benjegerdes et al.Psoriasis (Auckl) 2016; 6:131-44. Mahil et al. Semin Immunopathol 2016;38(1):11-27. Navarini et al. J Eur Acad Dermatol Venereol 2017;31(11):1792-9.). The generalized form (GPP) is associated withsignificant morbidity, and in some cases, mortality (Borges-Costa et al.Am J Clin Dermatol 2011; 12(4):271-6.). Loss-of-function mutations inthe IL-36RN gene are common in GPP patients, especially those withoutconcomitant symptoms of plaque psoriasis (Marrakchi et al. N Engl J Med2011; 365(7):620-8.). The IL-36RN gene encodes a protein known as theIL-36 receptor antagonist (IL-36RA). In healthy individuals, IL-36RAcompetes with activated IL36α, β and/or γ cytokines for binding to theIL36 receptor (IL-36R), giving this protein anti-inflammatory propertiesthat help maintain homeostasis. Aberrant structure and function ofIL-36RA engenders dysregulated secretion of inflammatory cytokines andchemokines (Marrakchi et al. N Engl J Med 2011; 365(7):620-8.). Here wereport that a small molecule antagonist of CCR6/CXCR2 was at moreeffective than anti-IL-17 therapy in reducing skin thickness andleukocyte infiltration after direct intradermal injection of activatedIL-36α into mouse skin.

GPP is characterized by a widespread eruption of pustules anderythematous plaques. In the acute variant, patients usually appearsystemically ill because the sudden eruption of pustules is accompaniedby pain and fever. Life threatening leukocytosis, electrolyteabnormalities, hypoalbuminemia and elevated liver enzymes can also occurin the acute variant (Benjegerdes et al. Psoriasis (Auckl) 2016;6:131-44). There are no approved treatments for GPP. Due to the rareoccurrence of this disease, there is no standardized method of assessingthe response to treatment. Data on treatment outcomes consist primarilyof retrospective studies, case reports and expert opinion (Benjegerdeset al. Psoriasis (Auckl) 2016; 6:131-44. Robinson et al. J Am AcadDermatol 2012; 67(2):279-88.).

Current treatments for GPP involve supportive care for those patientswho are systemically ill, followed by medical treatments to control theskin disease (Robinson et al. J Am Acad Dermatol 2012; 67(2):279-88.).Chronic, slowly progressing disease is typically managed by oralretinoids or methotrexate. Acute disease is treated with cyclosporine orone of the anti-TNFα biologics. None of these treatments are highlyeffective, and all have side effects (Robinson et al. J Am Acad Dermatol2012; 67 (2):279-88.). Thus, there is an unmet clinical need for newtreatments and new clinical targets for GPP management.

In addition to the T cells and neutrophils that accumulate in IL-36treated skin, this model also generates large numbers of myeloid cellswe believe to be iDC. This latter population expresses markers of bothDCs and macrophages. Since these cells do not express FLT3/CD135, webelieve them to be derived from monocytes rather than classical DC. Ithas been previously demonstrated that monocyte-derived DC from humanblood express high levels of IL-36R (Foster et al. J Immunol 2014;192(12):6053-61.). If the iDC from mouse skin prove to have similarproperties to human blood monocyte-derived DC, these cells may beimportant modulators of IL-36-induced inflammation in the model used inthe present study.

We used the IL36α model to compare the effectiveness of Compound 1.136to that of an α-IL-17RA MAb. Biologics against the IL-17 axis are oftenused as a second-line treatment for GPP in the clinic (Imafuku et al. JDermatol 2016; 43(9):1011-7.). We found the 90 mg/kg dose of Compound1.136 to be significantly more effective than α-IL-17RA (500mg/mouse/day) at reducing ear swelling (FIG. 8 ). Although α-IL-17RAtreatments significantly reduced the accumulation of neutrophils and iDCin skin, the CD4⁺ αβ T cell count was not affected. Thus, the additionalanti-inflammatory effect of Compound 1.136 over and above that ofα-IL-17RA closely corresponds to the reduction in CD4⁺ αβ T cells.

In summary, we have shown that the inflammation resulting from activatedIL-36α skin injections involves neutrophil, iDC and CD4 T cellaccumulation, similar to what is seen in GPP patients. Selectiveinhibition of CCR6 and CXCR2 by Compound 1.136 reduced all three ofthese inflammatory cell types and ameliorated skin inflammation.Blockade of the IL-17 axis reversed neutrophil and iDC but not CD4 Tcell accumulation in skin. Compound 1.136 is a more effectivetherapeutic agent than the saturating concentrations of an α-IL-17RAmonoclonal antibody assessed in this IL-36α induced model of psoriasis.These findings suggest that CCR6/CXCR2 antagonism may constitute a noveltarget class, and a mechanistically distinct therapeutic approach totreating dysregulation of the IL-36 cytokine axis (as in GPP), byspecifically acting upon the inflammatory cells that likely mediate thedisease.

Although the foregoing invention has been described in some detail byway of illustration and example for purposes of clarity ofunderstanding, one of skill in the art will appreciate that certainchanges and modifications may be practiced within the scope of theappended claims. In addition, each reference provided herein isincorporated by reference in its entirety to the same extent as if eachreference was individually incorporated by reference. Where a conflictexists between the instant application and a reference provided herein,the instant application shall dominate.

What is claimed is:
 1. A method of treating generalized pustularpsoriasis (GPP) in a subject in need thereof, said method comprisingadministering to the subject an effective amount of a compound ofFormula (A1)

wherein R^(1a) is selected from CH₃ and Cl; R^(1b) is H or CH₃; R³ is Hor D; R⁴ is H; R^(5a) and R^(5b) are each independently selected from H,F, Cl, Br and CH₃; R^(6a) and R^(6b) are each independently selectedfrom H and CH₃; and R⁷ is methyl or ethyl; or a pharmaceuticallyacceptable salt, solvate or hydrate, thereof.
 2. The method of claim 1,wherein R^(1a) is CH₃.
 3. The method of claim 1, wherein R^(1a) is Cl.4. The method of claim 1, wherein R^(1b) is H.
 5. The method of claim 1,wherein R^(1b) is CH₃.
 6. The method of claim 1, wherein R^(1a) andR^(1b) are both CH₃.
 7. The method of claim 1, wherein R³ is H.
 8. Themethod of claim 1, wherein R^(5a) is Cl, and R^(5b) is H.
 9. The methodof claim 1, wherein R^(5a) is H, and R^(5b) is F.
 10. The method ofclaim 1, wherein R^(6a) and R^(6b) are both H.
 11. The method of claim1, wherein R^(6a) is H, and R^(6b) is CH₃.
 12. The method of claim 1,wherein R⁷ is methyl.
 13. The method of claim 1, wherein R⁷ is ethyl.14. The method of claim 1, wherein R^(5a) Cl; R^(5b) is H; R^(6a) andR^(6b) are both H; and R⁷ is methyl.
 15. The method of claim 1, whereinR^(5a) is H; R^(5b) is F; R^(6a) is H; R^(6b) is CH₃; and R⁷ is methylor ethyl.